Many undesirable biological processes occur via ligand binding to cell surface receptors. Thus, it is sometimes desirable to have compounds and methods to reduce or modulate such binding.
The TGF-β superfamily includes a number of ligands of biological significance.
TGF-β and Activin play critical pathogenic roles in many diseases including the progression of cancer and uncontrolled fibrosis and scarring of tissues, e.g. kidney, lung and liver fibrotic diseases. Furthermore, Myostatin/GDF8 is another ligand which is related to Activin and which shares binding to the same Type II receptor (ActivinRIIb). Myostatin is a powerful inhibitor of skeletal muscle growth and is a validated therapeutic target for muscle wasting diseases such as muscular dystrophy. Bone morphogenetic proteins (BMP), which are other ligands in the TGF-β family, have been implicated in cardiovascular diseases. For example, high levels of both BMP2 and BMP4 have been found in calcified atherosclerotic plaques and diseased aortic valves.
Principal agents that target these ligands are ligand traps/antagonists that bind and sequester ligand. Two examples are: 1) anti-ligand antibodies and 2) soluble receptor ectodomains.
Efforts have been made to identify methods to reduce ligand binding by trapping ligand and preventing its interaction with the cell surface receptors. Inhibition of certain ligands has been reported using anti-ligand antibodies that trap and neutralize the ligand directly. For therapeutic and diagnostic applications, however, antibodies are problematic, particularly due to issues arising from their immunogenicity (and the danger of adverse immune response in patients) and their large size (restricting their ability to reach targets outside the bloodstream).
Soluble versions of receptor ectodomains antagonize ligands directly by binding to them and preventing them from interacting with cell surface receptors. In the case of TGF-β, in animal models, expression of a TGF-β receptor type II (TβRII) ectodomain (ED) partially restored host immunity and promoted tumor clearance, indicating that receptor ectodomain—mediated neutralization of TGF-β inhibits tumor progression. It has been shown, however, that the efficacy of monovalent TβRII to antagonize TGF-β is less than could be desired. Attempts to overcome this led to the production of an artificially dimerized form of versions of TβRII-ED, dimerized, via fusion to either coiled-coil domains or the Fc domain of IgG. This dimerization improved the antagonist effect.
Bivalent receptor-based traps/neutralizers that antagonize multimeric ligand activity have the potential to act as therapeutic or diagnostic (imaging or non-imaging) agents for diseases/disorders caused by over-production/activity of the target ligand. It has been demonstrated that non-covalent dimerization of TβRII-ED (for example, via fusion to heterodimerizing coil strands (coiled-coil TβRII-ED)), greatly enhances the antagonist potency of TβRII-ED (De Crescenzo et al., 2004, J. Biol. Chem. 279: 26013). A significant disadvantage of the coiled-coil fused dimer is that the non-covalent nature of the dimerization domain limits its potency, i.e. it dissociates at low concentrations such that a large portion of the coil-fused receptor ectodomain will be acting as a monomer rather than a dimer. Use of the Fc domain of IgG provides a covalent interaction, but at the cost of large size and increased probability of immunogenicity.